As the nervous system develops, a complex series of temporally and spatially regulated molecular events must occur to allow for the correct wiring of axons. The commissural axons of the spinal cord, which eventually develop into many of the sensory tracts in the adult, have served as a fundamental model for understanding paradigms of axon guidance. During vertebrate development these axons pathfind toward their eventual targets in a programmed fashion. As part of their trajectory, they must cross over the midline of the spinal cord, forming horizontal axon commissures. Once across, these axons turn and travel in a longitudinal plane toward their eventual target and do not recross the midline. The ability of an axon to grow towards its target is dependent on repulsive and attractive cues. These cues are mediated by a variety of newly identified receptor-ligand systems . The molecular events that attract axons toward the midline are at least partially understood, while the events that occur at the midline and thereafter are not. Recent work in Drosophila and C. elegans has identified a receptor that appears to control midline axon guidance events. Two rodent homologues of this receptor, termed Robo-1 and Robo-2, have been identified. Given their extensive sequence homology to the Drosophila and C. elegans Robo orthologues, and the fact that other basic axon guidance mechanisms are conserved across species, the Robo genes are strong candidate midline axon guidance molecules in vertebrates. The long term aim of this proposal is to illuminate the mechanisms through which growth cones make complex decisions. The specific focus of this project is the set of complex decisions that commissural growth cones make at the midline of the vertebrate nervous system, in particular, the role played by the two candidate receptors, Robo-1 and Robo-2 in directing midline axon guidance. The creation of Robo-deficient mice, and the identification of Robo ligands, the basis of this grant, should provide evidence supporting or disproving the role of these molecules in midline axon guidance. Clearly of basic interest, this area of research also has clinical ramifications with respect to understanding many of the neurodevelopmental genetic syndromes that exist, treating spinal cord injury, and developing means by which we might further understand the process of neuronal plasticity.